Membrane connectivity estimated by digital image analysis of HER2 immunohistochemistry is concordant with visual scoring and fluorescence in situ hybridization results: algorithm evaluation on breast cancer tissue microarrays

Abstract

IntroductionThe human epidermal growth factor receptor 2 (HER2) is an established biomarker for management of patients with breast cancer. While conventional testing of HER2 protein expression is based on semi-quantitative visual scoring of the immunohistochemistry (IHC) result, efforts to reduce inter-observer variation and to produce continuous estimates of the IHC data are potentiated by digital image analysis technologies.MethodsHER2 IHC was performed on the tissue microarrays (TMAs) of 195 patients with an early ductal carcinoma of the breast. Digital images of the IHC slides were obtained by Aperio ScanScope GL Slide Scanner. Membrane connectivity algorithm (HER2-CONNECT™, Visiopharm) was used for digital image analysis (DA). A pathologist evaluated the images on the screen twice (visual evaluations: VE1 and VE2). HER2 fluorescence in situ hybridization (FISH) was performed on the corresponding sections of the TMAs. The agreement between the IHC HER2 scores, obtained by VE1, VE2, and DA was tested for individual TMA spots and patient's maximum TMA spot values (VE1max, VE2max, DAmax). The latter were compared with the FISH data. Correlation of the continuous variable of the membrane connectivity estimate with the FISH data was tested.ResultsThe pathologist intra-observer agreement (VE1 and VE2) on HER2 IHC score was almost perfect: kappa 0.91 (by spot) and 0.88 (by patient). The agreement between visual evaluation and digital image analysis was almost perfect at the spot level (kappa 0.86 and 0.87, with VE1 and VE2 respectively) and at the patient level (kappa 0.80 and 0.86, with VE1max and VE2max, respectively). The DA was more accurate than VE in detection of FISH-positive patients by recruiting 3 or 2 additional FISH-positive patients to the IHC score 2+ category from the IHC 0/1+ category by VE1max or VE2max, respectively. The DA continuous variable of the membrane connectivity correlated with the FISH data (HER2 and CEP17 copy numbers, and HER2/CEP17 ratio).ConclusionHER2 IHC digital image analysis based on membrane connectivity estimate was in almost perfect agreement with the visual evaluation of the pathologist and more accurate in detection of HER2 FISH-positive patients. Most immediate benefit of integrating the DA algorithm into the routine pathology HER2 testing may be obtained by alerting/reassuring pathologists of potentially misinterpreted IHC 0/1+ versus 2+ cases.

Digital image analysis outperforms manual biomarker assessment in breast cancer

“…Individual applications for each biomarker then run the scoring of positive and negative cells itself, with subcellular resolution 40 ( Figure 1). Excellent reproducibility with this and similar systems has been shown previously [41][42][43][44][45] (see specific statistics on reproducibility with the Visiopharm integrator system for each tested biomarker in Supplementary Data).…”

High agreement between whole slide imaging and optical microscopy for assessment of HER2 expression in breast cancer

“…We also noted that WSI evaluation led to higher sensitivity of the antibodies than OM evaluation, and OM evaluation led to higher specificity of the antibodies. Laurinaviciene et al [21] described an almost perfect intraobserver agreement between visual evaluation and digital image analysis at the spot level of a TMA digitized slide (k = 0.86 and 0.87 in two visualizations). They used maximum TMA spot values (instead of mean or median value), which increases the sensitivity of HER2 detection.…”

Commentary

“…For instance, a recent position paper advocates that accurate assessment of common morphological end points in lung (e.g., alveolar number or size) requires the application of 3-D stereology (Hsia et al 2010). However, while mandated use of 3-D measurements may be justified in some settings, several recent studies across various disease indications in humans have shown that 2-D tIA of digital images yields data sets that are (1) concordant with scores acquired by conventional manual evaluation and diagnostic scoring and (2) at times provide an even more accurate measurement for the severity and stage of a disease (Daunoravicius et al 2014;Zhong et al 2016;Laurinaviciene et al 2011;Turashvili et al 2009;Vennalaganti et al 2015;Lee et al 2013;Farris et al 2014;Snead et al 2016;Ohlschlegel et al 2013). The utility of 2-D tIA gains additional importance in tissue specimens harvested in the course of human clinical trials because 2-D digital imaging approaches still provide useful information when tissue sample sizes are too small (e.g., needle core biopsies) to permit 3-D evaluation or when ethical considerations preclude optimal processing of tissue (i.e., euthanasia with rapid removal and fixation of tumor-ridden organs as practiced in animal experiments is not possible in human patients).…”